Abstract

2349

In contrast to protein antigens, processing of glycoproteins by dendritic cells (DC) for presentation to CD4+ or CD8+ T cells has not been studied. We have addressed this subject using as a model glycoprotein MUC1, a tumor antigen, expressed by the majority of human adenocarcinomas. DC can endocytose large amounts of soluble antigens for processing in the MHC class II pathway. Our published results show that DC efficiently endocytose soluble MUC1 glycopeptides, transport them to acidic compartments, process them into smaller peptides and present them on MHC class II molecules without removing the carbohydrates (Vlad et al, 2002, J. Exp. Med. 196: 1435). Exogenous soluble antigen can also access the cytosol, for processing in the MHC class I pathway (phenomenon known as cross-priming/cross-presentation). Processing of glycoproteins in the MHC class I pathway by DC has not been studied to date. We have investigated the fate of sugars on MUC1 glycopeptides that have been internalized by DC and have entered the cytosol for cross-priming. In order to increase the efficiency of antigen loading into the cytosol of DC, we coupled MUC1 to BioPORTER, a liposome-like protein delivery reagent. The complexed antigen can access both processing pathways and can trigger responses in both CD4+ and CD8+ T cell compartments. T cells primed on unglycosylated MUC1 do not cross-react when tested on DC pulsed with glycosylated MUC1, suggesting that DC do not deglycosylate the antigen and that glycopeptides are presented in MHC class I molecules as well. This preliminary finding is being confirmed through sequencing by ESI(Qtof)MS/MS of peptide fractions acid-eluted from the cell surface of glycosylated MUC1-loaded DC. MHC class I and II-restricted glycopeptides that are presented by DC provide an opportunity to elicit a much broader repertoire of T cells than can be expected solely based on the MUC1 peptide sequence. Immunotherapy with a MUC1 glycopeptide vaccine based on aberrantly glycosylated tumor MUC1 processed and presented by DC is currently under investigation in MUC1 transgenic mice.